Tissue-specific (or adult) stem cells are regarded as the source for normal tissue homeostasis and tissue repair. They also provide tremendous promise for regenerative medicine due to their capacity to proliferate and differentiate into a variety of mature cell types. Human mesenchymal stem cells (hMSCs) can differentiate into osteocytes, adipocytes, chondrocytes, muscle cells and neurons. However, the molecular mechanisms underlying these differentiating processes are poorly understood.
RNA-mediated interference (RNAi) is a highly conserved gene silencing event functioning through targeted destruction of individual mRNA by a homologous double-stranded small interfering RNA (siRNA) (Fire, A. et al., Nature 391:806-811 (1998)). siRNAs generated by both chemical synthesis and in vitro or in vivo transcription through vector-based expression systems have been proven very useful tools in studying gene loss-of-function in mammalian cells (Brummelkamp, T. R. et al., Science 296:550-553 (2002); Caplen, N. J. et al., Proc Natl Acad Sci USA 98:9742-9747 (2001); Elbashir, S. M. et al., Nature 411:494-498 (2001); Lee, N. S. et al., Nat Biotechnol 20:500-505 (2002); Miyagishi, P M. et al., Nat Biotechnol 20: 497-500 (2002); Paddison, P. J. et al., Genes Dev 16:948-958 (2002); Paul, C. P. et al., Nat Biotechnol 20:505-508 (2002); Sui, G. et al., Proc Natl Acad Sci USA 99:5515-5520 (2002); Yu, J. Y. et al., Proc Natl Acad Sci USA 99:6047-6052 (2002)). While high throughput screens using genome-scale siRNA libraries have been successfully carried out in mammalian cells (Berns, K. et al., Nature 428:431-437 (2004); Paddison, P. J. et al., Nature 428:427-431 (2004); Zheng, L. et al., Proc Natl Acad Sci USA 101:135-140 (2004)), effective application of arrayed synthetic siRNA library in stem cells has not been reported. Human mesenchymal stem cells (hMSCs) can be easily isolated from adults and expanded rapidly in vitro. Due to their ability to differentiate into various mature cell types (Prockop, D. J. Nat Biotechnol 20:791-792 (2002); Ryan, J. M. et al., J Inflamm (Lond) 2:8 (2005)) (Campagnoli, C. et al., Blood 98:2396-2402 (2001); Dezawa, M. et al., Science 309:314-317 (2005); Dezawa, M. et al., J Clin Invest 113:1701-1710 (2004); Pittenger, M. F. et al., Science 284:143-147 (1999); Pittenger, M. F. et al., Circ Res 95:9-20 (2004); Woodbury, D. et al., J Neurosci Res 61:364-370 (2000)), they have been of great interests to researchers exploring cell-based therapies for degenerative diseases including bone disorders (Dezawa, M. et al., Science 309:314-317 (2005); Dezawa, M. et al., J Clin Invest 113:1701-1710 (2004); Horwitz, E. M. et al., Proc Natl Acad Sci USA 99:8932-8937 (2002); Pittenger, M. F. et al., Circ Res 95:9-20 (2004); Prockop, D. J. Science 276:71-74 (1997)). Cell fate transition from stem cell self-renewal to differentiation involves not only positive regulators but also negative regulators that normally suppress differentiation.
There is a need for methodologies that influence the differentiation path of mesenchymal stem cells, for example, into cells of an osteoblast lineage. The present invention addresses this and other needs.